New chemicals are constantly produced either for consumer use or as by-products into the environment. These potential human carcinogens are tested in cultures of prokaryotes or lower eukaryotes, in living rodents and in mammalian cells in tissue culture. Although these tests are reproducible, reliable, quick, relatively inexpensive and do not sacrifice higher animals, they are inadequate for testing human carcinogens.
Current assays rely on mutagenicity or genotoxicity to identify carcinogens because historically, only mutagens were believed to be carcinogens. Nevertheless, not all carcinogens are mutagens. Mutagenic carcinogens are usually electrophiles or capable of metabolic conversion to electrophiles which attack DNA causing base alteration and mutation. Nonmutagenic carcinogens induce cell proliferation and DNA synthesis by a variety of biochemical mechanisms eventually resulting in genome alteration; but they are not initially mutagenic. Some metal cations such as vanadate act as mitogens or alter protein phosphorylation. The International Agency for Research on Cancer (IARC) has identified twenty-three chemicals and groups of chemicals which are causally associated with cancer in humans. Of these, arsenic compounds, diethylstilbestrol and others are only weakly mutagenic.
Cell transformation assays can detect both mutagenic and nonmutagenic carcinogens. Therefore, presumably, a chemical that induces or promotes transformation is a carcinogen or tumor promoter. To investigate chemical carcinogenesis and mechanisms or transformation, several assays have been developed which rely on cell transformation. (See, e.g., DiPaolo, J. A. et al. (1969) "Quantitative Studies of in vitro Transformation by Chemical Carcinogens," J. Natl. Cancer Inst., 42:867; Reznikoff, C. A. et al. (1973) "Establishment and Characterization of a Cloned Line of C3H Mouse Embryo Cells Sensitive to Postconfluence Inhibition of Cell Division," Cancer Res. 33:3231; Kakunaga, T. (1973) "A Quantitative System for Assay of Malignant Transformation by Chemical Carcinogens Using a Clone Derived from BALB/c3T3, " Intl. J. Cancer, 12:463). Transformed foci are the endpoint in these assays.
These tests, however, suffer from lack of reproducibility from laboratory to laboratory, technical difficulties, and difficulties in scoring foci as there are several different types of foci. Due to the low transformation frequency, large numbers of plates must be used to obtain statistically significant results for weak carcinogens. The C3H/10T1/2 assay requires six or more weeks of incubation before foci can be scored, increasing the loss of data due to contamination. Assays using cell transformation, nevertheless, respond to nongenotoxic carcinogens such as hormones and metals.
All virally-enhanced transformation assays have similar problems which compromise their usefulness. These include: variation in transformation frequency among various lots of cells, differing sensitivity to transfection and to carcinogens, variable rates of spontaneous transformation, technical complexity and limited commercial availability. Therefore, there exists a need for improved transformation assays and cell lines for use in such assays for rapid and reliable screening for carcinogens.